Unit

UNIT 14.15 Nonradioactive Analysis of Dynamic Protein Palmitoylation

  1. Brent R. Martin

Published Online: 24 SEP 2013

DOI: 10.1002/0471140864.ps1415s73

Current Protocols in Protein Science

Current Protocols in Protein Science

How to Cite

Martin, B. R. 2013. Nonradioactive Analysis of Dynamic Protein Palmitoylation. Current Protocols in Protein Science. 73:14.15:14.15.1–14.15.9.

Author Information

  1. Department of Chemistry, University of Michigan, Ann Arbor, Michigan

Publication History

  1. Published Online: 24 SEP 2013

Abstract

Methods to study protein S-palmitoylation dynamics have previously relied on metabolic labeling with [14C]palmitate, which requires additional safety precautions and long exposures. Nonradioactive alkynyl palmitate analogs have been developed for in-gel fluorescence detection and affinity purification. Cells metabolically labeled with the commercially available analog 17-octadynoic acid are lysed and then combined with azide-linked reporter tags for efficient conjugation by copper-catalyzed click chemistry in phosphate buffer. This approach has been demonstrated to label hundreds of endogenous palmitoylated proteins and is compatible with traditional pulse-chase methods. This protocol describes the reagents and procedures for labeling and detection of dynamic palmitoylation in mammalian cells. Curr. Protoc. Protein Sci. 73:14.15.1-14.15.9. © 2013 by John Wiley & Sons, Inc.

Keywords:

  • palmitoylation;
  • click chemistry;
  • post-translational modifications